Kang Y J, Cheng J, Mei L J, Hu J, Piao Z, Yin S X
Agricultural College, Yangzhou University, Yangzhou, China.
Mikrobiologiia. 2010 Sep-Oct;79(5):664-71.
The use of 16S rRNA gene has been a "golden" method to determine the diversity of microbial communities in environmental samples, phylogenetic relationships of prokaryotes and taxonomic position of newly isolated organisms. However due to the presence of multiple heterogeneous 16S rRNA gene copies in many strains, the interpretation of microbial ecology via 16S rRNA sequences is complicated. Purpose of present paper is to demonstrate the extent to which the multiple heterogeneous 16S rRNA gene copies affect RFLP patterns and DGG E profiles by using the genome database. In present genome database, there are 782 bacterial strains in total whose genomes have been completely sequenced and annotated. Among the total strains, 639 strains (82%) possess multiple 16S rRNA gene copies, 415 strains (53%) whose multiple copies are heterogeneous in sequences as revealed by alignment, 236 strains (30%) whose multiple copies show different restrict patterns by CSP61 + Hinfl, MspI + Rsal or HhaI as analyzed in silico. Polymorphisms of the multiple copies in certain strains were further characterized by G + C% and phylogentic distances based on the sequences of V3 region, which are linked to DGGE patters. Polymorphisms of a few strains were shown as examples. Using artificial communities, it is demonstrated that the presence of multiple heterogeneous 16S rRNA gene copies potentially leads to over-estimation of the diversity of a community. It is suggested that care must be taken when interpreting 16S rRNA-based RFLP and DGGE data and profiling an environmental community.
16S rRNA基因的使用一直是确定环境样本中微生物群落多样性、原核生物系统发育关系以及新分离生物分类地位的“黄金”方法。然而，由于许多菌株中存在多个异质的16S rRNA基因拷贝，通过16S rRNA序列对微生物生态学进行解读变得复杂。本文的目的是利用基因组数据库来证明多个异质的16S rRNA基因拷贝对限制性片段长度多态性（RFLP）模式和变性梯度凝胶电泳（DGGE）图谱的影响程度。在目前的基因组数据库中，总共有782个细菌菌株的基因组已被完全测序和注释。在这些菌株中，639个菌株（82%）拥有多个16S rRNA基因拷贝，415个菌株（53%）的多个拷贝在序列上是异质的（通过比对显示），236个菌株（30%）的多个拷贝在计算机模拟分析中通过CSP61 + Hinfl、MspI + Rsal或HhaI显示出不同的限制性模式。基于V3区域序列的G + C%和系统发育距离进一步表征了某些菌株中多个拷贝的多态性，这些与DGGE图谱相关。展示了一些菌株多态性的实例。通过人工群落证明，多个异质的16S rRNA基因拷贝的存在可能导致对群落多样性的高估。建议在解释基于16S rRNA的RFLP和DGGE数据以及分析环境群落时必须谨慎。
